The invention relates to a process for preparing substituted cyclopentane derivatives represented by the formula (I) 
wherein R1, R2, R3, R4, X and Y are as described below, and pharmaceutically acceptable salts thereof.
The invention further relates to a process for purifying the compound of formula (Ia) and novel crystalline forms of the compound of formula (Ia). 
The compounds of formula (I) are neuraminidase inhibitors, useful for the treatment of influenza and other viral infections.
The compounds of formula (I) and method of making and using compounds of formula (I) are described in pending application PCT US 98/26871, filed Dec. 17, 1997.
The invention relates to a more efficient process of preparing the compounds of formula (I), more particularly a process which permits high yield isolation of the desired stereoisomer, without the need for chromatographic purification.
The present invention relates to a process for preparing a compound of formula (I) 
wherein
R1 is selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl and aralkyl;
R2 is selected from the group consisting of hydrogen; alkyl, cycloalkyl, halogen substituted alkyl, aryl and substituted aryl;
R3 and R4 are independently selected from the group consisting of hydrogen, alkyl, alkylene, cycloalkyl, aryl and substituted aryl, provided that at least one of R3 or R4 is alkyl;
X is NHC(xe2x95x90NH)NH2;
Y is selected from the group consisting of hydrogen, fluorine, hydroxy, OR5, OCOR5, NH2, NHCOR5 and NR5R6, where R5 is selected from alkyl, aryl, COR6 or COOR6; and where R6 is selected from hydrogen, alkyl, alkylene, cycloalkyl, aryl or substituted aryl;
and pharmaceutically acceptable salts thereof;
comprising the steps of: 
reacting a compound of formula (II), wherein W is an N-protecting group, with a nitrile oxide of formula (III) by cycloaddition, at a temperature which prevents uncontrolled decomposition of the compound of formula (III), to yield a mixture of two stereoisomers of two regioisomers of the formula (IV) and (IVxe2x80x2), respectively; 
reacting the mixture of the compounds of formula (IV) and (IVxe2x80x2) by hydrolyzing to the corresponding alkali metal salts of formula (V) and (Vxe2x80x2), where M+ represents an alkali metal cation or a tetra-alkyl-ammonium cation; reacting the resulting mixture of salts of formula (V) and (Vxe2x80x2) with a acid to form the corresponding acids of formula (VI) and (VIxe2x80x2); reacting the acids of formula (VI) and (VIxe2x80x2) with ammonia or an amine, to precipitate the corresponding salt of formula (VII), where Q+ represents NH4+ or substituted ammonium cation; reacting the salt of formula (VII) to yield the compound of formula (IVa); 
reducing the compound of formula (IVa) using sodium borohydride and NiCl2 to yield a mixture of two stereoisomers of the corresponding aminoalcohol of formula (IX); isolating the desired diastereomer of formula (IXa) by recrystallization from an organic solvent or alcohol/water mixture; 
reacting the aminoalcohol of formula (IXa) with a compound of formula (X) or a compound of formula (XI), where V is chlorine or bromine, to produce the corresponding compound of formula (XII); 
if in the compound of formula I Y is other than hydroxy in the configuration as set forth in formula (XII), the compound of formula (XII) is further converted to the compound of formula (XIIxe2x80x2) by known methods, for example as described in pending application PCT US 98/26871, filed Dec. 17, 1997.;
removing the N-protecting group from the compound of formula (XII) or (XIIxe2x80x2) to produce the corresponding amine of formula (XIII), or salt thereof; hydrolyzing the compound of formula (XIII) to the corresponding acid salt of formula (XIV); and reacting the compound of formula (XIV) with a guanylating agent, to produce the compound of formula (I).
For the compound of formula (XIII), if Y represents NH2 or NR5R6 where R6 is hydrogen, then Y is protected with a protecting group other than W, which protecting group is removed by known methods following the guanylation step.
In another aspect, the invention is directed to a recrystallization procedure for purification and formation of a stable crystalline form of the compound of formula (Ia) 
comprising the steps of dissolving the crude compound of formula (Ia) in a mixture of water/methanol at reflux temperature; and cooling to crystallize the pure compound of formula (Ia).
The process of this invention, as described herein, is advantageous over previously disclosed methods in that it does not require chromatographic separation or purification; it does not require the use of highly flammable or toxic solvents such as ether, CH2Cl2, and the like; and it results in high product yield and purity; making it suitable for large scale production.
The recrystallization as described above may be directed to produce either of two crystalline forms of the compound of formula (Ia), labelled Form A and Form B, or a mixture thereof.
A further aspect of the claimed invention is directed to novel crystalline structures of the compound of formula (Ia), more particularly Form A and Form B, characterized by their respective X-ray powder diffraction patterns.
Form A of the compound of formula (Ia) is more stable to changes in ambient relative humidity.
As used herein, the term xe2x80x9calkylxe2x80x9d whether used alone or as part of a substituent group, include straight and branched chains containing one to eight carbon atoms, preferably one to three carbon atoms. For example, alkyl radicals include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, t-butyl, pentyl and the like.
As used herein, the term xe2x80x9calkylenexe2x80x9d whether used alone or as part of a substituent group, include straight, branched or cyclic unsaturated hydrocarbon groups containing two to eight carbon atoms, preferably two to three carbon atoms. Examples of alkylene groups include vinyl, 1-propenyl, allyl, 2-propenyl, 2-methyl-1-propenyl, cyclopentenyl, and the like.
As used herein, unless otherwise noted, xe2x80x9ccycloalkylxe2x80x9d shall denote cyclic aliphatic groups containing three to eight carbon atoms in the ring, optionally substituted with alkyl groups typically having one to six carbon atoms. Usually one or two substituent groups are present. Suitable examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.
As used herein, unless otherwise noted, xe2x80x9carylxe2x80x9d shall refer to unsubstituted carbocyclic aromatic groups containing six to fourteen carbon atom, such as phenyl, naphthyl, and the like, preferably phenyl.
As used herein, unless otherwise noted, xe2x80x9caralkylxe2x80x9d shall mean any alkyl group substituted with an aryl group. Suitable examples of aralkyl groups include benzyl, phenylethyl, and the like.
As used herein, unless otherwise noted, substituents on the aryl and aralkyl groups are one or more, preferably one or two, of halogen.
As used herein, xe2x80x9chalogenxe2x80x9d shall mean chlorine, bromine, fluorine and iodine.
As used herein, xe2x80x9caminexe2x80x9d shall mean a primary, secondary or tertiary amine such as ethylamine, t-butylamine, dimethylamine, diethylamine, triethylamine, piperidine, morpholine, N-methyl morpholine, and the like.
As used herein, xe2x80x9cguanylating agentxe2x80x9d shall mean an agent which adds a formamidine group to an amine nitrogen. Suitable examples of a guanylating agent include formamidine sulfonic acid, S-methyl iso-thiourea, 1H-pyrazole-1-carboxamidine monohydrochloride, 1H-triazole-1-carboxamidine monohydrochloride, and the like. Preferably, the guanylating agent is 1H-pyrazole-1-carboxamidine monohydrochloride or 1H-triazole-1-carboxamidine monohydrochloride, more preferably 1H-triazole-1-carboxamidine monohydrochloride.
As used herein, unless otherwise noted, an N-protecting group shall mean any functional group that is bonded directly to the N and is capable of preventing reactions at the N. Such protecting groups are known in the art, for example as described in Protective Groups in Organic Chemistry, ed. J. F. W. McOmie, Plenum Press, 1973; and Protective Groups in Organic Synthesis, 3rd Edition, T. W. Greene and P. G. M. Wuts, John Wiley and Sons, 1999. The appropriate choice of an N-protecting group can be made by one skilled in the art.
As used herein, the notation xe2x80x9c*xe2x80x9d shall denote the presence of a stereogenic center.
Where the compounds of the invention have one stereogenic center, they exist as enantiomers. Where the compounds possess two or more stereogenic centers, they additionally exist as diastereomers. It is to be understood that all such isomers and mixtures thereof are encompassed within the scope of the present invention.
With reference to substituents, the term xe2x80x9cindependentlyxe2x80x9d means that when more than one of such substituents is possible, such substituents may be the same or different from each other.
As used herein, the term xe2x80x9ca temperature which prevents uncontrolled decompositionxe2x80x9d shall mean a temperature safely below the decomposition temperature. For example, when in the compound of formula (III), R3 and R4 are each ethyl, the temperature which prevents uncontrolled decomposition is xe2x89xa690xc2x0 C.
Pharmaceutically acceptable salts of the compounds of formula (I) include those derived from pharmaceutically acceptable, inorganic and organic acids and bases. Examples of suitable acids include hydrochloric, hydrobromic, sulfuric, nitric, perchloric, fumaric, maleic, phosphoric, glycolic, lactic, salicylic, succinic, p-toluenesulfonic, tartaric, acetic, citric, methanesulfonic, formic, benzoic, malonic, naphthalene-2-sulfonic, trifluoroacetic and benzenesulfonic acids.
Salts derived from appropriate bases include alkali metal cations such as sodium, potassium, and the like; and ammonium ions such as ammonium, alkyl ammonium, and the like.
In a preferred embodiment, the claimed invention is used in preparing compounds of the formula (I) wherein R1, R2, R3 and R4 are independently selected from hydrogen or C1-C4 alkyl, R5 is hydrogen and Y is OH. More preferably, the claimed invention is used to prepare the compound of formula (Ia). 
The present invention relates to a process for preparing a compound of formula (I) as illustrated in Scheme 1 below: 
wherein R1, R2, R3, R4, X, Y, Q, V and W are as described above.
More particularly, an N-protected compound of formula (II), a known compound or compound prepared by known methods, is added to a nitrile oxide of formula (III), a compound prepared in situ by known methods, by cycloaddition in an inert organic solvent such as toluene, benzene, xylene, cyclohexane and the like, at a temperature which prevents uncontrolled decomposition of the compound of formula (III), preferably at a temperature between about 40xc2x0 C. and about 90xc2x0 C., in the presence of a tertiary amine such as triethylamine, diisopropylethylamine, N-methylmorpholine, and the like, preferably triethylamine, to yield a mixture of two stereoisomers of two regioisomers of formula (IV) and (IVxe2x80x2)
The substituted nitrile oxide of formula (III) can be produced in situ by known methods, preferably by heating a suitably substituted chlorooxime (represented by the formula R3R4CHxe2x80x94C(Cl)xe2x95x90NOH). For example, as described in Curran, D. P. and Gothe, S. A., Tetrahedron, 1988, 44(13), 3945-52.
To selectively isolate the desired stereoisomer, that is the compound of formula (IVa), the mixture of (IV) and (IVxe2x80x2), present in the organic solvent, is hydrolyzed with a basic aqueous solution, such as sodium hydroxide, potassium hydroxide and the like, preferably sodium hydroxide, to produce a mixture of the corresponding alkali metal salts of formula (V) and (Vxe2x80x2), respectively. The aqueous phase (containing the alkali metal salts) is separated from the organic phase and mixed with a suitable solvent such as an organic ether, for example methyl tert-butyl ether; an organic ester, for example ethyl acetate; a hydrocarbon, for example toluene; a water immiscible ketone, for example methyl-isobutyl ketone; or a water immiscible alcohol, for example n-butanol; preferably the solvent is methyl tert-butyl ether. The resulting mixture is treated with an acid, such as acetic, hydrochloric, sulfuric, and the like, preferably hydrochloric, to a pH of about 3-5, preferably to a pH of about 4, to produce the corresponding acids of formula (VI) and (VIxe2x80x2). The organic phase is separated and treated with ammonia or an amine, preferably t-butyl amine, preferably at a temperature between about 15-30xc2x0 C., more preferably at room temperature (20-25xc2x0 C.), resulting in precipitation of the corresponding salt, a compound of formula (VII), where Q+ represents NH4+ or substituted ammonium cation.
Alternatively, the reaction mixture containing the compounds of formula (V) and (Vxe2x80x2) is acidified with an acid, such as acetic, hydrochloric, sulfuric, and the like, preferably hydrochloric, to a pH in the range of about 0.5-3, preferably to a pH in the range of about 1-2, to produce the corresponding compounds of formula (VI) and (VIxe2x80x2). The organic phase is then separated and treated with ammonia or an organic amine, preferably t-butylamine, preferably at a temperature between about 15-30xc2x0 C., more preferably at room temperature, to produce the corresponding compound of formula (VII) as a precipitate, wherein Q+ is as previously defined.
If in the compound of formula (II), R1 is hydrogen, then the compound of formula (II) is reacted with the compound of formula (III), under the above described conditions, to produce the compounds of formula (VI) and (VIxe2x80x2) directly.
The salt of formula (VII) is suspended in an alcohol of the formula R1OH, a compound of formula (VIII), preferably treated with a corresponding orthoester. The resulting suspension is treated with a strong non-aqueous acid such as gaseous HCl, gaseous HBr, perchloric acid, p-toluene sulfonic, and the like, preferably gaseous HCl, at a temperature xe2x89xa620xc2x0 C., resulting in formation of the compound of formula (IVa). If the N-protecting group on the compound of formula (VII) is t-butoxycarbonyl (Boc), the solution is further treated with di-tert-butyl dicarbonate (Boc anhydride) and a tertiary amine such as triethylamine, diisopropylethylamine, N-methylmorpholine, and the like, preferably triethylamine.
Alternatively, if the N-protecting group on the compound of formula (VII) is t-butoxycarbonyl (Boc), the compound of formula (VII) is suspended in an organic solvent such as acetone, 2-butanol, acetonitrile, and the like, preferably acetone, and then treated with a base such as potassium carbonate, sodium carbonate, and the like, or an alkali hydroxide such sodium hydroxide, potassium hydroxide, and the like, or a mixture thereof, preferably a mixture thereof such as a mixture of sodium hydroxide and sodium carbonate. Preferably, the resulting suspension is heated to remove the amine byproduct of Q+, and then treated with an alkylating agent such as an alkyl halide, dimethyl sulfate, and the like, preferably dimethyl sulfate, to yield the corresponding compound of formula (Iva).
The stereoisomer of formula (IVa) is reduced to the corresponding compound of formula (IX) using sodium borohydride, preferably in the range of about 2-3 molar equivalents, and nickel chloride, preferably in an amount equal to at least 1 molar equivalent, in an alcohol such as methanol, ethanol, and the like, preferably in an alcohol of formula (VIII), preferably at a temperature of less than 10xc2x0 C. The product is isolated by addition of sodium nitrite, ammonium hydroxide and ammonium chloride, resulting in precipitation of the corresponding aminoalcohol of formula (IX).
The desired diastereomer of the compound of formula (IXa) is isolated by recrystallization from an organic solvent such as toluene, ethyl acetate, and the like; or alcohol/water mixture such as methanol/water, ethanol/water, and the like; preferably toluene.
Alternatively, the desired diastereomer of formula (IXa) is isolated by dissolving the mixture of formula (IX) in a weak acidic aqueous solution such as aqueous hydrochloric acid, acetic acid, and the like, adjusting the pH of the solution to pH greater than 7 with ammonium hydroxide, and extracting with an organic solvent such as ethyl acetate. The compound of formula (IXa) is then isolated by known methods.
The aminoalcohol of formula (IXa) is reacted with a compound of formula (X) or a compound of formula (XI), preferably a compound of formula (X), preferably with 1 to 2 equivalents, more preferably about 1 equivalent, in an inert organic solvent such as toluene, ethyl acetate, and the like, preferably toluene, to yield the corresponding compound of formula (XII).
If in the compound of formula (I) Y is other than hydroxy in the configuration as set forth in formula (XII), the compound of formula (XII) is further converted to the compound of formula (XIIxe2x80x2) by known methods, for example as described in pending application PCT US 98/26871, filed Dec. 17, 1997;
The N-protecting group on the compound of formula (XII) or (XIIxe2x80x2) is removed by known methods, to produce the corresponding amine of formula (XIII) or (XIIIxe2x80x2), respectively, or salts thereof, which amines or salts thereof are present in their dissolved forms within the separated aqueous phase.
When the N-protecting group on the compound of formula (XII) or (XIIxe2x80x2) is Boc, the N-protecting group is removed by treating the compound of formula (XII) or (XIIxe2x80x2), present in an organic solvent, with an acid such as aqueous hydrochloric or aqueous hydrobromic, preferably aqueous hydrochloric, at a pH in the range of 0-3, preferably to a pH less than 1, more preferably to a pH 0, preferably at a temperature in the range of 0-20xc2x0 C., to produce the salt of the corresponding amine of formula (XIII) or (XIIIxe2x80x2), respectively.
To the separated aqueous phase containing the compound of formula (XIII) or (XIIIxe2x80x2) is added a base, such as aqueous sodium hydroxide, aqueous potassium hydroxide, and the like, to adjust the pH to between about 6-13, preferably to a pH of 12-13, preferably at a temperature of less than or equal to about 10xc2x0 C., resulting in the compound of formula (XIV) or (XIVxe2x80x2), respectively. To the solution containing the compound of formula (XIV) or (XIVxe2x80x2) is added a guanylating agent, at a pH in the range of 7.5-14, preferably in the range of 8-10, at a temperature in the range of 0-90xc2x0 C., preferably at a temperature in the range of 10-50xc2x0 C., to yield the compound of formula (Ib) or (I), respectively.
Preferably, the guanylating agent is 1H-pyrazole-1-carboxamidine monohydrochloride, added at a pH about 9.5, at a temperature in the range of 40-50xc2x0 C.; or 1H-triazole-1-carboxamidine monohydrochloride, added at a pH about 8.5, at a temperature in the range of 20-30xc2x0 C. More preferably the guanylating agent is 1H-triazole-1-carboxamidine monohydrochloride.
Alternatively, the compound of formula (XII) may be hydrolyzed prior to removal of the protecting group.
If in the compound of formula (I) R 1 is other than hydrogen, the separated aqueous phase containing the compound of formula (XIII) or (XIIIxe2x80x2) is treated directly with a guanylating agent, at a pH in the range of 7.5-14, preferably in the range of 8-10, at a temperature in the range of 0-90xc2x0 C., preferably at a temperature in the range of 10-50xc2x0 C., to yield the corresponding compound of formula (Ib) or (I), respectively.
The compound of formula (Ia) is purified by dissolving the crude product in a mixture of methanol:water, preferably in the range of 5:95 to 50:50, more preferably 10:90, at reflux temperature. Preferably, the solution is maintained at reflux until the boiling temperature reaches between about 85xc2x0-100xc2x0 C., more preferably 98-100xc2x0 C. The resulting solution is cooled to precipitate the purified product of formula (Ia).
Recrystallization of the compound of formula (Ia) as described above will yield one of two novel crystalline forms, Form A or Form B, or a mixture thereof. Form A is obtained by slow recrystallization from the water/methanol solution, where the volume of the solution is xe2x89xa77 mL/g, preferably  greater than 10 mL/g, preferably where initial precipitation occurs at xe2x89xa630xc2x0 C. Form B is obtained by more rapid recrystallization from the water/methanol solution, where the volume of solution is xe2x89xa67 mL/g, preferably where initial precipitation occurs at  greater than 30xc2x0 C. Form B or a mixture of Form A and Form B can be completely converted to Form A by stirring the solid in water, preferably at a temperature of xe2x89xa720xc2x0 C., more preferably at a temperature of about 70-80xc2x0 C., and preferably cooling to a temperature of about 0-5xc2x0 C.
Preferably, Form A is prepared by rapid recrystallization of the compound of formula (Ia) to produce Form B or a mixture of Form A and Form B, followed by complete conversion to Form A.
The novel crystalline forms of the compound of formula (Ia) may be characterized by their respective x-ray powder diffraction patterns utilizing a Philips PW3710 based powder diffractometer using CuK60  radiation and the following system conditions:
Form A of the compound of formula (Ia) appears as small, high density shiny crystals, may exist as a di- or tri-hydrate and may be characterized by its X-ray diffraction pattern:
Form A of the compound of formula (Ia) may be characterized by an X-ray diffraction pattern comprising the following major peaks:
Form B of the compound of formula (Ia) appears as fluffy, low density needles, exists as a di-hydrate and may be characterized by its X-ray diffraction pattern:
Form B of the compound of formula (Ia) may be characterized by an X-ray diffraction pattern comprising the following major peaks:
The following examples describe the invention in greater detail and are intended to illustrate the invention, but not to limit it.